ChIP-seq experiments were analyzed both qualitatively and quantitatively. Reads were trimmed with the FASTX-Toolkit (-Q33 -t 20 -l 22), aligned with Bowtie 1.0 (-v 2 -m 1) on the Human hg38 reference genome, and peaks were called using MACS 2.1.1. H3K4me1, H3K27ac, H3K4me3, and H3K27me3 peaks were called with --broad using default parameters and q < 0.05.

Qualitative analysis, including intersection and comparison of bed files, was performed using BedTools version 2.23.

To define enhancer regions, we intersected those marked by H3K4me1 and H3K27ac in at least two samples, discarded regions with H3K4me3 in at least two samples, and discarded regions overlapping with TSS. Motif enrichment was performed by using HOMER v4.10.

Quantification of reads per region was performed with DeepTools 3.0.2. Differential mark deposition was conducted by means of edgeR 3.24.1 inside R 3.3.3. To define mark deposition following BAZ1B levels, we used the same design as for RNA-seq data (~individual+BAZ1B).

To identify BAZ1B bound regions and to avoid losing identification of lowly covered regions, we resorted to (i) aggregation of all sample aligned reads and (ii) peak calling with MACS2 using –extsize 800 and q < 0.25. BAZ1B binding coverage was calculated with DeepTools, with the same parameters used for histone marks, on the identified peak regions. Differentially bound regions were identified with edgeR.

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.